Rudolf Schipp

Electron microscopical and histochemical studies of differentiation and function of the cephalopod gill (Sepia officinalis L.)

SummaryPhase contrast and electron microscopical investigations on the gill ofSepia officinalis L., from the stage shortly before hatching to the adult stage, demonstrate (1) peripheral respiratory epithelial areas and (2) highly folded epithelia rich in mitochondria in the concave recesses of the gill lamellae. Enzyme-histochemical and cytochemical findings, and in particular the GOT proof, suggest by analogy with the chloride cells of the teleosts that the cells of this second type of epithelium are capable of active transport, and that they probably secrete ammonia. Apparently the gill of cephalopods, too, serves both respiratory and excretory functions.

Electron microscopic and histochemical studies on the function of the branchial heart appendage (pericardial gland) in Sepia officinalis (L.)

Die elektronenmikroskopischen Untersuchungen am Kiemenherzanhang von Sepia officinalis zeigen, das diesem Organ wie den Venenanhangen exkretorische Funktionen zukommen. Die Pericardialdruse erweist sich als ein Derivat des Kiemenherzens. Ihre Hauptmasse besteht aus einem polar organisierten Faltenepithel, dessen Ultrastrukturmerkmale (basaler Faltenapparat, Gehalt an Mitochondrien und Lysosomen, sekretorisch aktiver Mikrovillisaum) anderen Exkretionsorganen entsprechend auf aktive Ionentransportvorgange hindeuten. Es ist dem auseren Kiemenherzepithel homolog und kommt in ahnlicher Ausbildung auch in anderen Organen (Zentralherz, Kieme) vor. Die polygonalen Zellen der Randzone sind dem zentralen Gewebe des Kiemenherzens zu homologisieren; ihr Reusensystem in Nachbarschaft der Blutlakunen ist ahnlich dem der Podocyten in der Vertebratenniere und macht eine Druckfiltration wahrscheinlich. Als Filtermembranen funktionieren neben dem Plasmalemm auch noch die Basalmembran und Zwischenmembran, die porenartige, fur Myofer durchlassige Lucken von 30 A Weite zeigen. Daneben werden an der Grenze zwischen Epithel und Lakune wie auch in der Intima groserer Gefase besondere Dichtezellen dargestellt, die ahnlich wie Macrophagen grosere Partikel (z.B. Ferritin und Myofer) aufnehmen und intraplasmatisch als “dense bodies” ablagern.SummaryOur electron microscopic studies of the appendages of the branchial heart and veins of Sepia officinalis suggest an excretory function for these organs. The pericardial gland appears to be a derivate of the branchial heart; it mostly consists of a folded polarized epithelium. Ultrastructural features of the epithelium such as the basal folding system, contents of mitochondria and lysosomes, and secretory microvilli suggest active ion transport in these cells of the pericardial gland. The pericardial gland epithelium is homologous with the external epithelium of the branchial heart; it is also present in a similar form in the central heart and branchia. Polygonal cells present in the peripheral zone of the pericardial gland are homologous with the central branchial heart tissue. The system of basal cell processes of the polygonal cells near blood lacunae is similar to the podocyte-capillary relationship in the vertebrate kidney and suggests the possibility of pressure filtration. The plasmalemma, as well as the basal and intermediate membranes, appear to be filtration membranes; they contain pore-like structures about 30 Å in diameter which are permeable to Myofer. Furthermore, special dense cells are present at the epithelium-lacunae border and in the intima of larger vessels; these cells, similar to macrophages, take up large particles, such as ferritin and Myofer, and deposit them in the cytoplasm as dense bodies.Histochemical findings on the distribution of enzymes G-6-PD, SDH, IDH, MDH, and MAO, and the distribution in the folded epithelium and lacunae of alkaline and acid phosphatases confirm the ultrastructural results and suggest an excretory function of the pericardial gland.ZusammenfassungDie elektronenmikroskopischen Untersuchungen am Kiemenherzanhang von Sepia officinalis zeigen, daß diesem Organ wie den Venenanhängen exkretorische Funktionen zukommen. Die Pericardialdrüse erweist sich als ein Derivat des Kiemenherzens. Ihre Hauptmasse besteht aus einem polar organisierten Faltenepithel, dessen Ultrastrukturmerkmale (basaler Faltenapparat, Gehalt an Mitochondrien und Lysosomen, sekretorisch aktiver Mikrovillisaum) anderen Exkretionsorganen entsprechend auf aktive Ionentransportvorgänge hindeuten. Es ist dem äußeren Kiemenherzepithel homolog und kommt in ähnlicher Ausbildung auch in anderen Organen (Zentralherz, Kieme) vor. Die polygonalen Zellen der Randzone sind dem zentralen Gewebe des Kiemenherzens zu homologisieren; ihr Reusensystem in Nachbarschaft der Blutlakunen ist ähnlich dem der Podocyten in der Vertebratenniere und macht eine Druckfiltration wahrscheinlich. Als Filtermembranen funktionieren neben dem Plasmalemm auch noch die Basalmembran und Zwischenmembran, die porenartige, für Myofer durchlässige Lücken von 30 Å Weite zeigen. Daneben werden an der Grenze zwischen Epithel und Lakune wie auch in der Intima größerer Gefäße besondere Dichtezellen dargestellt, die ähnlich wie Macrophagen größere Partikel (z.B. Ferritin und Myofer) aufnehmen und intraplasmatisch als “dense bodies” ablagern.Die enzymhistochemischen Befunde über die Verteilung von G-6-PD, SDH, IDH, MDH und MAO und der sauren und alkalischen Phosphatase in Faltenepithel und Lakunen stützen diese Befunde und sprechen ebenfalls für eine exkretorische Funktion.

Components of the cellular defense and detoxification system of the common cuttlefish Sepia officinalis (Mollusca, Cephalopoda).

Endocytotic-active cells in the branchial heart complex of Sepia officinalis were studied by in situ injection of different types of xenobiotics and by in vitro perfusion of the organ complex with a bacterial suspension. The rhogocytes (ovoid cells) ingest particles of all tested sizes by endocytosis and phagocytosis. The hemocytes of the circulating blood and the adhesive hemocytes in the wall of the branchial heart incorporate all tested kinds of foreign materials, including bacterial cells due to phagocytosis achieved by the triangular mesenchymatic cells. The ultrastructural findings also give strong evidence that the triangular mesenchymatic cells are fixed hemocytes that have migrated into the branchial heart tissue. The ingestion and digestion of allogeneic substances and bacteria or their debris by rhogocytes and/or all (forms of) hemocytes suggests the involvement of these either fixed or mobile endocytotic-active cells in the defense and detoxification system of cephalopods.

Cytomorphology and function of the pericardial appendages of Nautilus (Cephalopoda, Tetrabranchiata)

SummaryThe pericardial appendages (glands) are the principal excretory organs of the tetrabranchiate cephalopods. They were studied in Nautilus macromphalus and N. pompilius using light and electron microscopical methods. The four organs, which are homologous with the branchial heart complex in Coleoida, appear to represent a phylogenetically archaic state. Actually, they have had to take over the primary functions of secretion, and it will be seen that they are highly evolved. They are built up of numerous contractile villi which can be subdivided into three functionally important areas: the folded peripheral epithelium functioning particularly in reabsorption, the primarily secretory epithelium of the apical infoldings of the villi, and the ovoid cells with foot processes building up an ultra-filtration barrier. The filtrate seems to be expelled into the terminal secretory epithelium of the apical infoldings and not directly into the coelom.Within the brush border of the peripheral epithelia there are commensal flagellated bacteria, which may occupy an ecological niche similar to that of Dicyemida in the renal sacs of coleoids.

Ultrastructural and cytochemical investigations on the renal appendages and their concrements in dibranchiate cephalopods (Mollusea, Cephalopoda)

SummaryThe renal appendages of the octopods Octopus vulgaris Lam., Eledone moschata Lam. and of the decapods of the order Sepioidea Sepia officinalis L., and of sea water suggest osmotic filtration combined with active ion transport and methods of light and electron microscopy and of cytochemistry. The cells of the excretory epithelium are polarised; in all species, they show a basal labyrinth with a thick positive PAS and alcian blue reaction. The cells contain numerous mitochondria, which show the crista type of internal structure, and in the basal as well as in the apical part there are lysosomal dense bodies with high phosphatase activity. The fine structure, the osmolality and the ionic composition of the blood, the urine and of sea water suggest osmotic filtration combined with active ion transport and apocrine excretory secretion and reabsorption. In Sepia only, crypts of the epithelium were found to contain spherical mixed crystals which contain Ca, K, Na, Mg, Cl, S, P according to the electron probe and cytochemical analysis; the matrix shows concentric layers, it gives a positive PAS and alcian blue reaction. The chemical composition, the genesis and the possible function of these crystals are discussed.

Cytology of the renal appendages of Nautilus (Cephalopoda, Tetrabranchiata).

SummaryThe renal appendages of tetrabranchiate cephalopods, Nautilus pompilius (L.) and Nautilus macromphalus (G.B. Sow.), were studied using light- and electron-microscopical methods. The appendages, homologous to the renal appendages of dibranchiate cephalopods, possess in both species a folded transporting epithelium characterized by a secreting brush border and a high content of mitochondria and lysosome-like dense bodies as well as a basal labyrinth, and are separated from the blood lacunae by a multilayered lamina basalis.In the extracellular crypt-like infoldings of the epithelium, numerous concentric stratified concrements are found.The results strengthen the hypothesis that these concrements represent mineral-storage structures only found in calcium shell-supporting species within the class Cephalopoda.

Localization of catecholamine-containing nerve fibres in the branchial heart and cardiac ganglion of the common cuttlefish Sepia officinalis L. (cephalopoda)

The innervation of the branchial heart of Sepia officinalis was examined using TEM and glyoxylic acid induced fluorescence. In the cardiac ganglion and in cardiac nerves bluish-green fluorophores were seen associated with perikarya and varicose nerve fibres. Microspectrofluorometric analysis provided clear evidence that monoaminergic neurons in the branchial heart contain only catecholamines. Considering pharmacological data, it is more than likely that 5-hydroxytryptamine (serotonin) is not present in this system.

Cytomorphology and copper content of the basal cells in the midgut gland of Nautilus (Cephalopoda, Tetrabranchiata)

SummaryBy means of light and electron microscopical studies on Nautilus pompilius and Nautilus macromphalus, special basal cells within the alveolary enlarged terminal parts of the tubules of the midgut gland were revealed. Their abundant rough endoplasmic reticulum (RER) with enlarged cisterns synthesizes ring-shaped macromolecules that resemble the hemocyanin molecules within the bloods-paces in size and form and seem to be secreted throughout the lamina basalis, locally discharged into the adjacent blood sinus. The hypothesis that this cell type represents a site of hemocyanin synthesis was confirmed by Zeeman AAS and ASTEM analyses of the content and distribution of copper within the glandular tissue, especially the different compartments of this cell type.

Distribution and function of biogenic amines in the heart of Nautilus pompilius L. (Cephalopoda, Tetrabranchiata)

SummaryBiogenic amines (serotonin and catecholamines), play an important role in the control of the blood flow not only in vertebrates, but also in invertebrates such as cephalopods. In contrast to the well investigated hearts of the ȁ8modern,ȁ9 coleoid cephalopods, the innervation of the heart of the archaic Nautilus pompilius L. has not been studied in detail. In this study the distribution and effects of biogenic amines in the Nautilus heart were investigated. Serotonin and catecholamines were visualised by the glyxoylic acid induced fluorescence. High performance liquid chromatotography analysis was performed to discriminate between the catecholamines, which showed a high content of noradrenaline in the 4 auricles, the aorta and the ventricle, whereas the ventricle showed a high dopamine content. Adrenaline was found at a very low concentration in the ventricle. Serotonin and dopamine were also immunohistochemically localised to larger nerves and throughout the heart, respectively. In organ bath experiments, the auricles showed little spontaneous activity. After adding serotonin, they displayed rhythmical contractions, which were accelerated dose-dependently by noradrenaline. In summary, these data suggest an important role for biogenic amines in the control of the heart of Nautilus pompilius L., with serotonin possibly stimulating excitatory nerve fibres, whereas noradrenaline is likely to influence the muscle contraction itself.

Localization of putative neurotransmitters in the mantle and siphuncle of the mollusc Nautilus pompilius L. (Cephalopoda)

The neurotransmitter supply in the nerve endings of the mantle and the siphuncle, i.e. in organs that are responsible for the shell formation in the ectocholeate Nautilus pompilius, were investigated with electron microscopical, fluorescence-, immuno- and enzyme histochemical methods as well as with high pressure liquid chromatography (HPLC). Using antibodies against serotonin and the tetrapeptide FMRF-amide, positive reactions were demonstrated immunohistochemically within the terminal nerve fibres of the mantle and the vessels of the siphuncle. Enzyme histochemical proof of the presence of specific acetylcholinesterase yielded positive results in the muscle fibres of the mantle and siphuncle. Additionally, in the mantle, glyoxylic acid-induced fluorescence was shown within the nerve endings indicating catecholamines as neurotransmitters, whereas in the siphuncle such fluorescence did not appear. However, the HPLC-analyses showed that in the mantle and also in the siphuncle the content of dopamine is higher than that of noradrenaline whereas only traces of adrenaline occur in both organs suggesting dopamine as a putative neurotransmitter. Transmission electron microscopical examination of the nerve endings of both organs revealed that different types of vesicles were distinguished that could be considered as cholinergic, aminergic and peptidergic structures.